Melatonin effect on platelet count in patients with liver disease

Aim: A positive effect of melatonin on platelet count in patients with chronic liver disease is reported in the current study. Background: Thrombocytopenia occurs when the severity of liver disease is exacerbated. Reduction in the secretion of thrombopoetin, as an intrinsic hormone produced mainly by the liver, plays an important role in this complication induced by liver disease. Methods: This research was a double-blind, cross-over, placebo-controlled pilot study. Patients with liver disease were given two 5-mg pearls of melatonin or a placebo for two weeks, and after a 2-week washout period, their groups were switched. Liver function tests and platelet counts were assessed once at the beginning and once at the end of each phase of the study. Results In the current study, 40 patients meeting the eligibility criteria were included. The average platelet count was significantly increased by melatonin in comparison with the placebo (from 175.67±92.84 to 191.10±98.82 vs. from 185.22±98.39 to 176.45±91.45) (p-value <0.001). Melatonin also significantly reduced ALT, AST, total bilirubin, and direct and MELD scores in patients with liver disease (p-value <0.05). Conclusion: Melatonin may increase platelet count and inhibit thrombocytopenia in patients with liver disease; however, more investigations are needed to confirm the current results.


Introduction
1 Thrombocytopenia, as one of the complications caused by chronic liver disease (CLD), can be defined as a platelet count less than 150000 per μL (1). The prevalence rate of thrombocytopenia ranges widely from 6% to 78% in patients, depending on the type of liver disease (2)(3)(4). The liver plays a main role in the production and destruction of platelets. As reported in previous studies, there is a direct correlation between the severity of thrombocytopenia and the stage of liver disease (5). Some mechanisms for thrombocytopenia induced by liver disease have been explained, such as a decrease in Thrombopoietin as an endogenous hormone released by liver, which plays a main role in this complication (6). A greater reduction in thrombopoietin production occurs when liver disease is exacerbated, like in a higher stage of fibrosis (7). Other causes of thrombocytopenia in liver disease include suppression of the bone marrow production of platelets due to inflammation and platelets being trapped in the spleen (1). Melatonin is an endocrinic hormone secreted by different parts of body, such as the liver and the pineal BRIEF REPORT gland, into the gastrointestinal system and plasma, respectively (8). Different therapeutic effects for melatonin have been observed, such as anti-oxidant and hepato-protective effects, regulation of the circadian rhythm, and the amelioration of side effects induced by chemotherapy (9). Moreover, the Thrombopoietic property of melatonin has been demonstrated in previous human studies (10,11). In their study, Lissoni et al. used 20 mg of melatonin as adjuvant therapy during chemotherapy in patients with cancer and reported a reduced prevalence of myelosuppression and thrombocytopenia (10). According to pharmacological studies, melatonin is effective on thrombocytopenia associated with some cytokines such as IL-2, IL-12, TNF, and interferon alpha and stimulated megakaryocytes fragmentation into platelets (12,11). Because of the lack of proper pharmacological agents for the treatment of thrombocytopenia in chronic liver disease, further clinical investigations are needed to propose a pharmacological therapy with less unpleasant side effects. This report is a part of the results of a randomized clinical trial (no. IRCT20180519039718N1) in which the anti-pruritic effect of melatonin in liver disease was evaluated.

Methods
This cross-over, double-blind, randomized, placebocontrolled trial was conducted on patients with pruritus induced by liver disease who referred to the Liver Disease Clinic affiliated with Tehran University of Medical Sciences (TUMS) in Tehran, Iran, from July 15, 2018 to January 31, 2019.
The In this RCT, the included participants were allocated into two groups receiving either melatoninplacebo or placebo-melatonin by computer-generated randomization in four block sizes (A, B, C, and D).
The patients were allocated into two groups that received either two 5-mg pearls (along with 10 mg at night orally) of melatonin (NutraLab Company, Canada, supplied by Zahravi Pharmaceutical Companies, Iran) or 2 pearls of a placebo (Zahravi Pharmaceutical Companies, Iran) for a 2-week period. Then, after a two-week washout period, patient groups were switched. None of the patients had received any new medicine which could have increased platelet count in the 4 weeks prior to being enrolled in this study.
Liver function tests, including alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total and direct bilirubin, serum creatinine (SrCr), international normalized ratio (INR) for calculating "model for end-stage liver disease" (MELD) score, and complete blood count (CBC) with differentiation and platelet count were assessed once at the beginning and once at the end of each treatment duration (laboratory data was evaluated 4 times). Eligible patients who had experienced pruritus induced by liver disease for at least 4 weeks were included in this study. The exclusion criteria were any history of hypersensitivity to melatonin, uncontrolled epilepsy, pregnancy, breastfeeding, decompensated liver disease, unstable hemodynamic conditions such as a mean arterial pressure <65 mmHg (13), or a chronic kidney disease with creatinine clearance <15 ml/min or dialysis (14).

Statistical analysis
SPSS software (version 20.0, SPSS Inc., Chicago, IL, USA) was used for statistical analyses. Continuous data was reported as mean ± SD, non-normal values were presented by median (interquartile range), and categorical values were presented by frequency (percentage). To evaluate the normality distribution, the Kolmogorov-Smirnov test was performed on numerical variables. An independent t-test and Mann-Whitney U test were used to compare parametric and nonparametric variables, respectively. Cohen's d was calculated with a 95% confidence interval (95% CI) (15). A p-value < 0.05 was considered as statistically significant.

Results
In this study, 40 eligible patients were followed up until the end of the research. The demographic data of the patients are summarized in Table 1.
The platelet count was significantly elevated by melatonin compared to the placebo (p-value <0.05) ( Table 2). Notably, the effect size (Cohen's d) was 0.95 (CI 95%, 0.30_1.61). Moreover, 72.5% of 40 patients had an increase in platelet count, the average percent of which was 18.13% ± 18.74%, while 42.5% of 40 patients in the placebo group had an increase in their platelet count with an average of 7.45 ± 5.88 (p-value <0.05). Figures 1 and 2 show the trend of platelet count for each patient who received melatonin or the placebo, respectively.   Table 1). Participants received no drugs or supplements which could affect the platelet count.

Discussion
In this study, platelet count was significantly increased by melatonin consumption ( (16). In fact, according to a systematic review performed on the therapeutic effects of melatonin in cancer, melatonin may be considered for the treatment of chemotherapy-induced thrombocytopenia. Although the exact protective mechanism of melatonin against platelet destruction is still unknown, it is currently used as an adjuvant treatment during chemotherapy (16). Paolo et al. studied a heterogenic population in radiation oncology or hematology and showed the thrombopoietic properties of melatonin. Moreover, they showed that the platelet count of 84% of 32 patients, Abbreviations: Alanine transaminase (ALT), Aspartate transaminase (AST), Alkaline phosphatase (ALP), International normalized ratio (INR), Liver function tests (LFTs), Platelets (PLT), Serum Creatinine (SCr), MELD Score (Model For End-Stage Liver Disease). All data are presented in mean±SD. P-value <0.05 is significant. * p-value for comparing patients' difference of parameter after and before each phase exposure) † pvalue 0.03 due to negative effect. who had thrombocytopenia related to either cirrhosis (7 patients) or liver metastases (25 patients) and then received 20 mg of melatonin for one month, reached the normal range of at least 100000 /μL, and the average percentage of increase in platelets was estimated to be 185%± 34% (11). The duration of treatment and dose of melatonin were both higher in Paolo's study than in the current one; therefore, it seems that the higher dose and longer duration more efficiently improved platelet count. The response to thrombopoietic therapy is defined as an increase by more than 30000/ μL or doubling of the baseline count (17); however, in studies on liver disease-induced thrombocytopenia, such as Kalambokis et al., the thrombopoietic properties of rifaximin 1200 mg for one month in cirrhotic patients were reported as inhibitors of bacterial overgrowth. It was also shown that rifaximin significantly increased platelet counts in comparison with a placebo (88900 ± 37200 / μL to 109700± 39700/ μL vs. 76800 ± 30300/ μL to 81100± 28500 / μL, p-value <0.05) (18). The current, study demonstrated that melatonin decreased ALT (84.32±78.11 to 59.20±37.86), AST (82.37±60.44 to 68.57±44.21), direct bilirubin (3.14±5.31 to 2.52±4.71), total bilirubin (5.23±10.03 to 4.22±7.59), and MELD score (11.17±5.62 to 9.77±5.27), all of which were statistically significant (p-value <0.05). The current results reflected the findings of Chojnacki et al.'s placebo-controlled study, confirming that consumption of 5 mg of melatonin twice daily significantly decreased the elevated liver enzymes that were induced by statins (p-value <0.001) (19). Some studies have also demonstrated that melatonin has some hepato-protective properties (20,19). Herein, it was hypothesized that alleviation of the severity of liver disease by the anti-fibrotic and antioxidant properties of melatonin may lead to the increased production of thrombopoietin by the liver. Melatonin significantly increased platelet counts in patients with liver disease compared to the placebo (pvalue < 0.05). This study was not designed to evaluate the thrombopoietic properties of melatonin; therefore, it would be worthwhile to perform studies to determine the optimum dose of melatonin, onset of action, and duration of the thrombopoietic properties.